This plot displays Total Ion Chromatograms (TICs) derived from MS1 scans across all analyzed samples. The x-axis represents retention time, and the y-axis shows the total ion intensity at each time point. Each colored trace corresponds to a different sample. The TIC provides a global view of the ion signal throughout the LC-MS/MS run, reflecting when compounds elute from the chromatography column. Key aspects to assess include:

• Overall intensity pattern: A consistent baseline and similar peak profiles across samples indicate good reproducibility.
• Major peak alignment: Prominent peaks appearing at similar retention times suggest stable chromatographic performance.
• Signal-to-noise ratio: High peaks relative to baseline noise reflect better sensitivity.
• Chromatographic resolution: Sharp, well-separated peaks indicate effective separation.
• Signal drift: A gradual decline in signal intensity across the run may point to source contamination or chromatography issues.

Deviations such as shifted retention times, missing peaks, or inconsistent intensities may signal problems in sample preparation, LC conditions, or mass spectrometer performance that require further investigation.

The Base Peak Chromatogram (BPC) displays the intensity of the most abundant ion at each retention time point across your LC-MS run. Unlike the Total Ion Chromatogram (TIC) which shows the summed intensity of all ions, the BPC highlights the strongest signals, providing better visualization of compounds with high abundance while reducing baseline noise. This makes it particularly useful for identifying major components in complex samples, monitoring dominant species, and providing clearer peak visualization when signal-to-noise ratio is a concern. Comparing BPC patterns across samples allows you to evaluate consistency in the detection of high-abundance compounds and can reveal significant variations in sample composition or instrument performance.

This plot shows the number of peaks detected in MS1 scans over the course of each sample run. The x-axis represents retention time (in minutes), while the y-axis displays the number of distinct ion signals (peaks) identified in each MS1 scan. The MS1 peak count reflects spectral complexity and provides insight into instrument performance during the LC-MS analysis. Key aspects to consider include:

• Overall pattern: Peak counts typically increase during the elution of complex mixtures and decrease during column washing or re-equilibration phases.
• Peak density: Higher counts suggest more complex spectra, potentially indicating a greater number of compounds present at that time point."
• Peak Consistency across samples: Similar profiles among replicates or related samples indicate good analytical reproducibility.
• Sudden drops: Abrupt decreases in peak count may point to transient ionization issues, spray instability, or chromatographic disruptions.
• Baseline values: The minimum peak count observed reflects the level of background noise or instrument sensitivity in the absence of eluting compounds.

Monitoring MS1 peak counts complements total ion chromatogram (TIC) and base peak chromatogram (BPC) data, offering an additional layer of quality control related to signal complexity, instrument stability, and sample composition.

This table presents general statistics for MS1 information extracted from mass spectrometry data files." It displays MS runs with their acquisition dates and times. For each file, the table shows two key metrics: TotalCurrent (the sum of all MS1 ion intensities throughout the run) and ScanCurrent (the sum of MS2 ion intensities). These values provide a quick overview of the total ion signals detected during both survey scans (MS1) and fragmentation scans (MS2), allowing for comparison of overall signal intensity across samples. Consistent TotalCurrent and ScanCurrent values across similar samples typically indicate good reproducibility in the mass spectrometry analysis, while significant variations may suggest issues with sample preparation, instrument performance, or ionization efficiency. The blue shading helps visualize the relative intensity differences between samples.

This plot shows the pipeline performance overview. Some metrics are calculated.

• Heatmap score[Contaminants]: as fraction of summed intensity with 0 = sample full of contaminants; 1 = no contaminants
• Heatmap score[Pep Intensity (>23.0)]: Linear scale of the median intensity reaching the threshold, i.e. reaching 2^21 of 2^23 gives score 0.25.
• Heatmap score[Charge]: Deviation of the charge 2 proportion from a representative Raw file (median). For typtic digests, peptides of charge 2 (one N-terminal and one at tryptic C-terminal R or K residue) should be dominant. Ionization issues (voltage?), in-source fragmentation, missed cleavages and buffer irregularities can cause a shift (see Bittremieux 2017, DOI: 10.1002/mas.21544 ).
• Heatmap score [MC]: the fraction (0% - 100%) of fully cleaved peptides per Raw file
• Heatmap score [MC Var]: each Raw file is scored for its deviation from the ‘average’ digestion state of the current study.
• Heatmap score [ID rate over RT]: Judge column occupancy over retention time. Ideally, the LC gradient is chosen such that the number of identifications (here, after FDR filtering) is uniform over time, to ensure consistent instrument duty cycles. Sharp peaks and uneven distribution of identifications over time indicate potential for LC gradient optimization.Scored using ‘Uniform’ scoring function. i.e. constant receives good score, extreme shapes are bad.
• Heatmap score [MS2 Oversampling]: The percentage of non-oversampled 3D-peaks. An oversampled 3D-peak is defined as a peak whose peptide ion (same sequence and same charge state) was identified by at least two distinct MS2 spectra in the same Raw file. For high complexity samples, oversampling of individual 3D-peaks automatically leads to undersampling or even omission of other 3D-peaks, reducing the number of identified peptides.
• Heatmap score [Pep Missing]: Linear scale of the fraction of missing peptides.

This table shows the quantms pipeline summary statistics.

Including Sample Name、Possible Study Variables、identified the number of peptide in the pipeline、 and identified the number of modified peptide in the pipeline, eg. All data in this table are obtained from the out_msstats file. You can also remove the decoy with the remove_decoy parameter.

This statistic is extracted from the out_msstats file. Proteins supported by more peptide identifications can constitute more confident results.

This information can be used to identify potential ionization problems including many 1+ charges from an ESI ionization source or an unexpected distribution of charges. MALDI experiments are expected to contain almost exclusively 1+ charged ions. An unexpected charge distribution may furthermore be caused by specific search engine parameter settings such as limiting the search to specific ion charges.

This chart assumes centroid data. Too few peaks can identify poor fragmentation or a detector fault, as opposed to a large number of peaks representing very noisy spectra. This chart is extensively dependent on the pre-processing steps performed to the spectra (centroiding, deconvolution, peak picking approach, etc).

Generally, one should expect to have a high number of low intensity noise peaks with a low number of high intensity signal peaks. A disproportionate number of high signal peaks may indicate heavy spectrum pre-filtering or potential experimental problems. In the case of data reuse this plot can be useful in identifying the requirement for pre-processing of the spectra prior to any downstream analysis. The quality of the identifications is not linked to this data as most search engines perform internal spectrum pre-processing before matching the spectra. Thus, the spectra reported are not necessarily pre-processed since the search engine may have applied the pre-processing step internally. This pre-processing is not necessarily reported in the experimental metadata.

For high complexity samples, oversampling of individual 3D-peaks automatically leads to undersampling or even omission of other 3D-peaks, reducing the number of identified peptides. Oversampling occurs in low-complexity samples or long LC gradients, as well as undersized dynamic exclusion windows for data independent acquisitions.

• Heatmap score [EVD: MS2 Oversampling]: The percentage of non-oversampled 3D-peaks.

Mass deltas close to zero reflect more accurate identifications and also that the reporting of the amino acid modifications and charges have been done accurately. This plot can highlight systematic bias if not centered on zero. Other distributions can reflect modifications not being reported properly. Also it is easy to see the different between the target and the decoys identifications.

The quantification information of peptides is obtained from the MSstats input file. The table shows the quantitative level and distribution of peptides in different study variables, run and peptiforms. The distribution show all the intensity values in a bar plot above and below the average intensity for all the fractions, runs and peptiforms.

• BestSearchScore: It is equal to 1 - min(Q.Value) for DIA datasets. Then it is equal to 1 - min(best_search_engine_score[1]), which is from best_search_engine_score[1] column in mzTab peptide table for DDA datasets.
• Average Intensity: Average intensity of each peptide sequence across all conditions with NA=0 or NA ignored.
• Peptide intensity in each condition (Eg. CT=Mixture;CN=UPS1;QY=0.1fmol): Summarize intensity of fractions, and then mean intensity in technical replicates/biological replicates separately.

The quantification information of proteins is obtained from the msstats input file. The table shows the quantitative level and distribution of proteins in different study variables and run.

• Peptides_Number: The number of peptides for each protein.
• Average Intensity: Average intensity of each protein across all conditions with NA=0 or NA ignored.
• Protein intensity in each condition (Eg. CT=Mixture;CN=UPS1;QY=0.1fmol): Summarize intensity of peptides.